From a plasmid (pBR322) clone bank of R strain Rickettsia rickettsii DNA, we have identified an Escherichia coli transformant expressing the 155-Kilodalton (Kd) major protective antigen of R. rickettsii. The transformant, EM24(pGAM21), harbors pBR322 with a 10.1-Kilosbase (Kb) insert of rickettsial DNA. A 3.7-Kb PstI fragment containing the antigen-encoding gene was subcloned downstream of the lactose promoter on pUC8 to generate pGAM22. Sonic lysates of E. coli harboring pGAM22 were used to sucessfully vaccinate mice against a lethal challenge of viable R. rickettsii and to vaccinate guinea pigs protecting them from Rocky Mountain spotted fever upon challenge with R. rickettsii.